SUNY Downstate Health Sciences University

Department of Physiology and Pharmacology

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Stanley Friedman, PhD

Repair of DNA Cross Links

We are studying the effect of DNA damage caused by DNA-protein cross links, the effect
of these cross links on cell viability and DNA replication and the pathways that repair
them.

The role of methylation reactions in the control of cellular processes especially
by the methylation of DNA, the effect of nucleoside analog inhibitors of DNA methylases
on cell growth, the mechanism of DNA repair as it relates to removal of DNA-protein
cross links.

We have been studying the properties of DNA methylases and of drugs that inhibit their
activity. Several of these proteins are autoregulated. In addition to binding to the
DNA at the site that they methylate, they bind to their promoter inhibiting transcription
of their respective genes. The properties of these binding reactions are very different.
The recognition sequence that confers specificity for methylation consists of only
5 nucleotides. However, when they bind to the promoter they protect 50bp from DNase
I digestion. One of our objectives is to determine the basis for the difference in
binding specificity of these proteins at these sites.

We have demonstrated that DNA methylases can be inhibited by nucleoside analogs. The
analog must be incorporated into DNA to be an effective inhibitor. These analogs cause
the DNA methylase to bind tightly to the DNA containing them. Some of these analogs
will kill the cells only when they contain DNA methylase; the cells are presumably
killed by the binding of the enzyme to DNA. The cells can repair this DNA damage.
They die when the repair system can no longer repair all the DNA-protein cross links
that are formed. We have been studying the repair of these DNA-protein adducts by
preparing DNA containing the analog in vitro and studying the survival of the DNA
in vivo. The cells can replicate the DNA-protein complex with approximately 50% of
the efficiency that they can replicate unbound DNA. Replication is not dependent on
either the recA or uvrA gene product. Presently we are studying processes by which
these DNA-protein adducts are repaired.